Preparation of Sample Support Films in Transmission Electron Microscopy using a Support Floatation Block
File(s)Preparation of Sample Support Films.pdf (776.33 KB)
Published version
Author(s)
Aylett, Christopher
Type
Journal Article
Abstract
Structure determination by cryo-electron microscopy (cryo-EM) has rapidly grown in the last decade; however, sample preparation remains a significant bottleneck. Macromolecular samples are ideally imaged directly from random orientations in a thin layer of vitreous ice. However, many samples are refractory to this, and protein denaturation at the air-water interface is a common problem. To overcome such issues, support films-including amorphous carbon, graphene, and graphene oxide-can be applied to the grid to provide a surface which samples can populate, reducing the probability of particles experiencing the deleterious effects of the air-water interface. The application of these delicate supports to grids, however, requires careful handling to prevent breakage, airborne contamination, or extensive washing and cleaning steps. A recent report describes the development of an easy-to-use floatation block that facilitates wetted transfer of support films directly to the sample. Use of the block minimizes the number of manual handling steps required, preserving the physical integrity of the support film, and the time over which hydrophobic contamination can accrue, ensuring that a thin film of ice can still be generated. This paper provides step-by-step protocols for the preparation of carbon, graphene, and graphene oxide supports for EM studies.
Date Issued
2021-04-08
Date Acceptance
2021-04-08
Citation
Journal of Visualized Experiments
ISSN
1940-087X
Publisher
MyJove Corporation
Start Page
1
End Page
11
Journal / Book Title
Journal of Visualized Experiments
Volume
170
Issue
1
Copyright Statement
Copyright © 2021 JoVE Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License.
Sponsor
Wellcome Trust
Identifier
https://www.jove.com/t/62321/preparation-sample-support-films-transmission-electron-microscopy
Grant Number
206212/Z/17/Z
Subjects
0601 Biochemistry and Cell Biology
1701 Psychology
1702 Cognitive Sciences
Publication Status
Published
Article Number
e62321
Date Publish Online
2021-04-08